PROJECT SUMMARY/ABSTRACT Roughly 85% of triple-negative breast cancers are categorized as basal-like or claudin-low carcinoma, molecular subtypes with especially poor prognosis and limited treatment options. Triple-negative breast cancers frequently harbor mutations in DNA-surveillance pathways; consequently, their overall genomic heterogeneity has been extensively characterized. By comparison, much less work has been done on the cell biology of triple-negative breast cancer. Despite the recognized histological nonuniformity of triple-negative tumors, we have only a rudimentary inventory of the types of signaling and transcriptional regulatory states that single basal-like and claudin-low cells can adopt. The long-term goal of this work is to identify and characterize the major cell-to-cell regulatory heterogeneities in triple-negative breast cancer. The current application focuses on growth-differentiation factor 11 (GDF11), a diffusible factor that is heterogeneously regulated in 3D organotypic cultures of claudin-low breast epithelial cells. Functional GDF11 bioactivity is lost in clinical cases of advanced triple-negative breast cancer, and addition of GDF11 to invasive claudin-low and basal-like cancer lines strongly suppresses invasion into basement membrane ECM. The hypothesis is that GDF11 acts a local breast-epithelial cue for proper lobular architecture, which is suppressed nongenetically during triple-negative breast cancer progression. The aims of this proposal are: 1) To identify the signaling and transcriptional mechanisms that mediate GDF11-induced phenotypes in triple-negative breast cancer. 2) To define the key steps of GDF11 misregulation in triple-negative neoplasms. 3) To determine the impact of GDF11 on progression and metastatic colonization of triple-negative tumors. The diversity of regulatory states enables triple-negative breast cancer cells to switch and adapt rapidly during tumor progression and the evolution of drug resistance. A complete inventory of regulatory states and their transitions could one day be harnessed by novel therapies that reset intratumor regulatory heterogeneity to delay progression or resistance.